Discharge volume control method, discharge pressure control method, and microbody forming method

ABSTRACT

A second valve is opened to combine an injection-pressure generating pressure with a maintaining pressure into an injection pressure. After the injection pressure is applied to a capillary to cause discharge of an object therefrom, an output pressure of a regulator is set to the injection pressure. Then, a first valve is opened to reapply the injection pressure to the capillary. The second valve is opened to combine a maintaining-pressure generating pressure with the injection pressure into the maintaining pressure. After the maintaining pressure is applied to the capillary to terminate the discharge of the object, the output pressure of the regulator is set to the maintaining pressure. Then, the first valve is opened to reapply the maintaining pressure to the capillary.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of and claims the benefit of U.S.application Ser. No. 11/785,431, filed Apr. 17, 2007, and claims benefitunder 35 U.S.C. Section 119 of Japanese Patent Application No.2006-270053, filed Sep. 29, 2006 the disclosures of all of which areincorporated herein by reference.

BACKGROUND

1. Field

The present invention relates to a discharge volume control method, adischarge pressure control method, an injecting apparatus, a microbodyforming method, a discharge volume control device, and a dischargevolume control program

2. Description of the Related Art

A microinjection, which is a method of injecting a specific substance(DNA, a drug, or the like) into a cell under a microscope, has beenknown in the fields of regenerative medicine, new drug designing, andthe like. Microinjection is a method of pressurizing a capillary filledwith a solution containing a predetermined substance in advance, therebydischarging the solution into a cell penetrated by the capillary toinject the substance into the cell. This method allows, for example, toexamine effects of the injected substance on the cell. A quantitativedischarge of a solution into a cell is critically important formicroinjection. Therefore, liquid discharge volume control methods forregulating a discharge volume conveniently and accurately with highrepeatability have been developed.

For example, Japanese Patent Application Laid-open No. H3-119989(Paragraphs 0010 to 0014, FIG. 1, and FIG. 2) discloses a microinjectionapparatus in which a male screw is rotated to move a plunger, therebypressurizing a capillary to discharge liquid. Specifically, when anoperator presses a button provided on a control box included in themicroinjection apparatus, an electric signal is generated, and thegenerated electric signal causes a piezoelectric element to produce adrive force to rotate the female screw so that the plunger is moved todischarge liquid filled in the capillary. Accordingly, the operator ofthe microinjection apparatus achieves the discharge of the liquid out ofthe capillary penetrated into a cell conveniently at the simple push ofthe button.

While the conventional technology described above achieves discharge ofliquid conveniently by moving a plunger with the push of a button, adischarge volume is regulated by an operator of the microinjectionapparatus who observes a cell penetrated by the capillary under amicroscope and estimates the discharge volume based on a degree ofexpansion of the cell to determine whether a predetermined liquid volumehas been discharged. Therefore, the conventional technology still has aproblem of not attaining a quantitative discharge of the liquid.

SUMMARY

It is an object of the present invention to at least partially solve theproblems in the conventional technology.

According to an aspect of the present invention, a discharge volumecontrol method for controlling a microinjection apparatus that includesa regulator regulating and outputting pressure, a capillary filled withan object to be discharged and connected to the regulator by a tube, afirst valve located on regulator side in the tube and a second valvelocated on capillary side in the tube, and controlling volume of theobject discharged from the capillary, includes opening the second valvefrom a state where the first valve and the second valve are closed tocombine an injection-pressure generating pressure with a maintainingpressure to generate an injection pressure, the injection-pressuregenerating pressure being retained between the first valve and thesecond valve to generate the injection pressure that causes the objectto be discharged from the capillary, and the maintaining pressure havingbeen applied to the capillary to prevent backflow of the object into thecapillary, applying the injection pressure to the capillary to causedischarge of the object from the capillary, setting an output pressureof the regulator to the injection pressure, opening the first valve toreapply the injection pressure to the capillary, opening the secondvalve from the state where the first valve and the second valve areclosed to combine a maintaining-pressure generating pressure with theinjection pressure having been applied to the capillary to generate themaintaining pressure, the maintaining-pressure generating pressure beingretained between the first valve and the second valve to generate themaintaining pressure, applying the maintaining pressure to the capillaryto terminate the discharge of the object, setting the output pressure ofthe regulator to the maintaining pressure, and opening the first valveto reapply the maintaining pressure to the capillary.

According to another aspect of the present invention, a dischargepressure control method for controlling pressure to discharge an objectfrom a capillary connected to a pressure regulator and injecting theobject into a microbody, includes closing a first valve that is locatedbetween the pressure regulator and the capillary in a state wherepressure in the capillary is maintained at a first pressure, thepressure regulator generating a second pressure, closing a second valvethat is located between the pressure regulator and the first valve,opening the first valve to combine the first pressure with the secondpressure to generate a third pressure, applying the third pressure tothe capillary to cause discharge of the object from the capillary, thepressure regulator resetting the third pressure, opening the secondvalve, closing the first valve, the pressure regulator generating afourth pressure, closing the second valve, and opening the first valveto maintain the pressure in the capillary at the first pressure.

According to still another aspect of the present invention, a dischargepressure control method for controlling pressure to discharge an objectfrom a capillary connected to a pressure regulator and injecting theobject includes closing a first valve that is located between thepressure regulator and the capillary in a state where pressure in thecapillary is maintained at a first pressure, the pressure regulatorgenerating a second pressure, closing a second valve that is locatedbetween the pressure regulator and the first valve, opening the firstvalve to combine the first pressure with the second pressure to generatea third pressure, applying the third pressure to the capillary to causedischarge of the object from the capillary, and the pressure regulatorresetting the third pressure in a state where the object has beendischarged from the capillary.

According to still another aspect of the present invention, a dischargepressure control method for controlling pressure to discharge an objectfrom a capillary connected to a pressure regulator, includes thepressure regulator generating a second pressure in a state wherepressure in the capillary is maintained at a first pressure, combiningthe first pressure with the second pressure to generate a thirdpressure, applying the third pressure to the capillary to causedischarge of the object from the capillary, the pressure regulatorresetting the third pressure while the object is being discharged fromthe capillary, and maintaining the pressure in the capillary at thethird pressure.

According to still another aspect of the present invention, a dischargepressure control method for controlling pressure to discharge an objectfrom a capillary connected to a pressure regulator, includes closing avalve that is located in a passage between the pressure regulator andthe capillary in a state where pressure in the capillary is maintainedat a predetermined pressure, measuring pressure in the passage betweenthe valve and the capillary with a pressure gauge that is locatedbetween the valve and the capillary, and indicating a result of themeasurement when the pressure falls below a predetermined thresholdvalue during a predetermined time period.

According to still another aspect of the present invention, an injectingapparatus that injects an object into a microbody, includes a capillarythat injects the object into the microbody, a regulator that regulatespressure to be applied to the object, a first valve that is located in apassage between the capillary and the regulator, a second valve that islocated between the first valve and the regulator, and a controller thatcontrols the regulator, the first valve, and the second valve. Thecontroller closes the first valve in a state where pressure in thecapillary is maintained at a first pressure, causes the regulator togenerate a second pressure, and closes the second valve. The controllerthen opens the first valve to combine the first pressure with the secondpressure to generate a third pressure, applies the third pressure to thecapillary to cause discharge of the object from the capillary, andcauses the regulator to reset the third pressure while the object isbeing discharged from the capillary.

According to still another aspect of the present invention, a microbodyforming method for forming a microbody into which a capillary injects anobject, includes inserting the capillary into the microbody in a statewhere pressure in the capillary is maintained at a first pressure, aregulator generating a second pressure, combining the first pressurewith the second pressure to generate a third pressure, applying thethird pressure to the capillary to inject the object from the capillaryinto the microbody, the regulator resetting the third pressure while theobject is being injected, maintaining the pressure in the capillary atthe third pressure, the regulator generating a fourth pressure,maintaining the pressure in the capillary at the first pressure byapplication of the third pressure and a fourth pressure, and removingthe capillary from the microbody.

According to still another aspect of the present invention, a dischargevolume control device that controls a microinjection apparatus thatincludes a regulator regulating and outputting pressure, a capillaryfilled with an object to be discharged and connected to the regulator bya tube, a first valve located on regulator side in the tube and a secondvalve located on capillary side in the tube, and controls volume of theobject discharged from the capillary, includes an injection pressureregulating unit that opens the second valve from a state where the firstvalve and the second valve are closed to combine an injection-pressuregenerating pressure with a maintaining pressure to generate an injectionpressure, the injection-pressure generating pressure being retainedbetween the first valve and the second valve to generate the injectionpressure that causes the object to be discharged from the capillary, andthe maintaining pressure having been applied to the capillary to preventbackflow of the object into the capillary, applies the injectionpressure to the capillary to cause discharge of the object from thecapillary, sets an output pressure of the regulator to the injectionpressure, and opens the first valve to reapply the injection pressure tothe capillary, and a maintaining pressure regulating unit that opens thesecond valve from a state where the first valve and the second valve areclosed to combine a maintaining-pressure generating pressure with theinjection pressure having been applied to the capillary to generate themaintaining pressure, the maintaining-pressure generating pressure beingretained between the first valve and the second valve to generate themaintaining pressure, applies the maintaining pressure to the capillaryto terminate the discharge of the object, sets the output pressure ofthe regulator to the maintaining pressure, and opens the first valve toreapply the maintaining pressure to the capillary.

According to still another aspect of the present invention, acomputer-readable recording medium stores therein a computer programthat causes a computer to implement the above method.

The above and other objects, features, advantages and technical andindustrial significance of this invention will be better understood byreading the following detailed description of presently preferredembodiments of the invention, when considered in connection with theaccompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a schematic of a microinjection apparatus according to a firstembodiment of the present invention;

FIGS. 2A to 2C are schematics for explaining an overview and features ofa liquid discharge volume control device shown in FIG. 1;

FIG. 3 is a block diagram of the liquid discharge volume control deviceshown in FIG. 1;

FIG. 4 is flowchart of a process performed by the liquid dischargevolume control device shown in FIG. 1;

FIG. 5 is a schematic for explaining an overview and features of aliquid discharge volume control device according to a second embodiment;

FIG. 6 is flowchart of a process performed by the liquid dischargevolume control device according to the second embodiment;

FIG. 7 is a schematic for explaining an overview and features of aliquid discharge volume control device according to a third embodimentof the present invention;

FIG. 8 is a block diagram of the liquid discharge volume control deviceaccording to the third embodiment;

FIG. 9 is a flowchart of a process performed by the liquid dischargevolume control device shown in FIG. 8;

FIG. 10 is a diagram of a computer that executes a computer program toimplement the liquid discharge volume control device shown in FIG. 1;

FIG. 11 is a schematic of a microinjection apparatus according to aconventional technology;

FIGS. 12A and 12B are schematics for explaining an overview and featuresof a liquid discharge volume control device shown in FIG. 11;

FIG. 13 is a schematic for explaining calculations of injection-pressuregenerating pressure and maintaining-pressure generating pressure; and

FIG. 14 is a schematic for explaining an accumulated error inconsecutive discharging.

DETAILED DESCRIPTION

Exemplary embodiments of the present invention are explained in detailbelow with reference to the accompanying drawings.

First, an example of a microinjection apparatus is explained. Asdisclosed in Japanese Patent Application Laid-Open No. 2006-133512,“METHOD FOR DISCHARGING LIQUID INTO CELL AND MICROINJECTION APPARATUS”by the inventor of this application, a liquid discharge volume controlmethod for regulating a discharge volume by switching between twopressures, i.e., a pressure (hereinafter, “injection pressure”) by whicha solution (object) is discharged into a cell (microbody) and a pressure(hereinafter, “maintaining pressure”) by which backflow to a capillaryis prevented, is available.

Specifically, in the conventional liquid discharge volume controlmethod, a liquid volume to be discharged out of the capillary into acell is quantitatively controlled by quickly generating an injectionpressure in a state where the capillary receives a maintaining pressureso that discharge of liquid out of the capillary is started, and, aftera lapse of a predetermined time, quickly reducing the pressure to themaintaining pressure to terminate the discharge of liquid out of thecapillary. The conventional liquid discharge volume control method isbriefly explained. FIG. 11 is a schematic of a microinjection apparatus1000 according to the conventional technology. The microinjectionapparatus 1000, which is connected to a positive pressure pump 1 and anegative pressure pump 2, includes a regulator 3, a regulating chamber5, a capillary 9, a valve a4, a valve b7, a pressure sensor a6, and apressure sensor b8. The regulator 3 maintains a pressure developed bythe pumps 1 and 2 constant. The regulating chamber 5 traps the pressuremaintained by the regulator 3. The capillary 9 is filled with asolution. The valve a4 is between the regulator 3 and the regulatingchamber 5. The valve b7 is between the regulating chamber 5 and thecapillary 9. The pressure sensor a6 detects a pressure in the regulatingchamber 5. The pressure sensor b8 detects a pressure applied to thecapillary. The regulator 3, the valve a4, the regulating chamber 5, thepressure sensor a6, the valve b7, the pressure sensor b8, and thecapillary 9 are connected by a tube 10 (indicated by thick lines). Aliquid-discharge-volume control device 20 that employs the liquiddischarge volume control method controls operations of themicroinjection apparatus 1000. For example, as each of the valves a4 andb7, a valve including an electromagnetic solenoid is employed. Theliquid-discharge-volume control device 20 generates an electric signal,thereby controlling opening and closing operations of each of the valvesa4 and b7.

In an actual configuration, rather than providing the regulating chamber5 in the microinjection apparatus 1000, a portion of the tube 10 (theportion where the valve a4, the pressure sensor a6, and the valve b7 areconnected), the portion indicated as an area surrounded by the dottedline in (B) of FIG. 11, forms the regulating chamber 5. Hereinafter, theregulating chamber 5 is the area surrounded by the dotted line in (B) ofFIG. 11.

FIG. 12A and FIG. 12B are schematics for explaining an overview andfeatures of the liquid discharge volume control device 20 according tothe conventional technology.

First, with reference to FIG. 12A, steps that cover until application ofan injection pressure Pi to the capillary 9 are explained. Theliquid-discharge-volume control device 20 causes the regulator 3 togenerate a maintaining pressure Pc by using the positive pressure pump 1and the negative pressure pump 2, and then opens the valves a4 and b7(see (1) of FIG. 12A). Subsequent to this step, an operator connects thecapillary 9 filled with a solution to the tube 10. Hence, the liquidfilled in the capillary 9 is retained in the capillary 9 without flowingbackward. The state shown in (1) of FIG. 12A is maintained untilpenetration of a cell with the capillary 9 by the operator under amicroscope to discharge the liquid becomes available.

When penetration of the cell with the capillary 9 by the operator tostart injection becomes available, as shown in (2) of FIG. 12A, theliquid-discharge-volume control device 20 closes the valve b7 tomaintain the capillary 9 under the maintaining pressure Pc in accordancewith an operator request to start injection, and causes the regulator 3to generate an injection-pressure generating pressure Ph (describedlater), which is a pressure necessary to generate the injection pressurePi, by using the positive pressure pump 1 and the negative pressure pump2 (see (3) of FIG. 12A).

As shown in (4) of FIG. 12A, the liquid-discharge-volume control device20 closes the valve a4 to maintain the regulating chamber 5 under thepressure Ph, and then opens the valve b7. Consequently, theinjection-pressure generating pressure Ph and the maintaining pressurePc are combined into the injection pressure Pi to start liquiddischarging (see (5) of FIG. 12A). As explained above, instantaneousopening/closing of the valves a4 and b7 allows to change the pressureapplied to the capillary from the maintaining pressure Pc to theinjection pressure Pi quickly.

Subsequently, with reference to FIG. 12B, steps that cover until theinjection pressure Pi is reduced to the maintaining pressure Pc toterminate the discharge of liquid out of the capillary are explained. Asshown in (6) of FIG. 12B, the liquid discharge volume control device 20closes the valve b7 to maintain the capillary 9 under the injectionpressure Pi, and opens the valve a4 (see (7) of FIG. 12B). Theliquid-discharge-volume control device 20 then causes the regulator 3 togenerate a maintaining-pressure generating pressure PL (describedlater), which is necessary to generate the maintaining pressure Pc, byusing the positive pressure pump 1 and the negative pressure pump 2 (see(8) of FIG. 12B).

As shown in (9) of FIG. 12B, the liquid discharge volume control device20 closes the valve a4 to maintain the regulating chamber 5 under themaintaining-pressure generating pressure PL, and then opens the valve b7(see (10) of FIG. 12B). Consequently, the maintaining-pressuregenerating pressure PL and the injection pressure Pi are combined intothe maintaining pressure Pc to terminate the discharge of liquid out ofthe capillary 9. As explained above, instantaneous opening/closing ofthe valves a4 and b7 allows to change the pressure applied to thecapillary from the injection pressure to the maintaining pressurequickly. Accordingly, the injection pressure that takes a rectangularwaveform can be applied to the capillary 9, which allows to performaccurate control of a liquid discharge volume by adjusting the injectionpressure and an injection-pressure application time (a period of timerequired to perform steps (5) of FIG. 12A to (10) of FIG. 12B).

When the liquid-discharge-volume control device 20 is required toconsecutively discharge liquid to another cell, theliquid-discharge-volume control device 20 causes the regulator 3 togenerate the maintaining pressure Pc, and opens the valve a4 to shiftthe state from that shown in (10) of FIG. 12B to that shown in (1) ofFIG. 12A, to perform operation control including the same steps asabove. As explained above, even when injection is to be performedconsecutively, it is possible to penetrate the capillary 9 into anothercell and to shift to the injection stage under a pressure that isquickly reduced to the maintaining pressure Pc. Therefore, the liquiddischarge volume can be accurately controlled also in the subsequentcycle.

The above-mentioned injection-pressure generating pressure Ph forgenerating the injection pressure Pi and the maintaining-pressuregenerating pressure PL for generating the maintaining pressure Pc areexplained. A relation between a pressure at a point in time before thevalve b7 is opened and a pressure at a point in time after the valve b7is opened can be obtained using an equation of state of gas. As shown in(B) of FIG. 11, the gas volume on the capillary 9 side is denoted as V1,the pressure on the capillary 9 side at a point in time before the valveb7 is opened as P1, the gas volume of the regulating chamber 5 as V2,and the gas volume of the regulating chamber 5 at a point in time beforethe valve b7 is opened as P2. In this case, relations between thepressure P, which is a pressure at a point in time after the valve b7 isopened, and a volume ratio η (η=V2/N1) are expressed by each ofequations (1) and (2) of FIG. 13. Accordingly, a pressure P2 that needto be retained in the regulating chamber 5 before the valve b7 is openedto generate the pressure P after the valve b7 is opened can be expressedby equation (3) of FIG. 13.

Meanwhile, a pressure P1 on the capillary 9 side at a point in timebefore the valve b7 is opened can be obtained using the pressure sensorb8, the pressure P2 in the regulating chamber 5 at a point in timebefore the valve b7 is opened can be obtained using the pressure sensora6, and the pressure P at a point in time after the valve b7 is openedcan be obtained using the pressure sensors b7 and a6. Therefore, thevolume ratio η can be obtained in advance from equation (2) of FIG. 13using the pressure values determined by performing a preliminarydischarge of liquid.

The injection-pressure generating pressure Ph or themaintaining-pressure generating pressure PL is calculated using thevolume ratio η and equation (3) of FIG. 13. The injection-pressuregenerating pressure Ph is calculated by substituting the injectionpressure Pi for the pressure P, which is the pressure at a point in timeafter the valve b7 is opened, and the maintaining pressure Pc for thepressure P1, which is the pressure on the capillary 9 side at a point intime before the valve b7 is opened, in equation (3) of FIG. 13. Themaintaining-pressure generating pressure PL is calculated bysubstituting the maintaining pressure Pc for the pressure P, which isthe pressure at a point in time after the valve b7 is opened, and theinjection pressure Pi for the pressure P1, which is the pressure on thecapillary 9 side at a point in time before the valve b7 is opened, inequation (3) of FIG. 13. The liquid-discharge-volume control device 20causes the regulator 3 to generate the injection-pressure generatingpressure Ph at step (3) of FIG. 12A, and the maintaining-pressuregenerating pressure PL at step (8) of FIG. 12B.

However, the above-explained method has a problem in that, even when theinjection-pressure generating pressure Ph or the maintaining-pressuregenerating pressure PL is calculated, a change in the “volume ratio η”caused by elements such as flexibility of the material of the tube 10,pressure values, and a pressure-applying time develops a slight error inthe target pressure in relation to at least one of an actually generatedinjection-pressure generating pressure Ph or an actually generatedmaintaining-pressure generating pressure PL, which results in unstableaccuracy of the liquid discharge volume.

The above-explained method has another problem in that, because an errorin an output of a regulator that regulates the pressure trapped in theregulating chamber 5 is not considered, when liquid is to beconsecutively discharged on a cell-by-cell basis, errors in outputs areaccumulated, which results in unstable accuracy of the liquid dischargevolume. Put another way, in consecutive discharging, when an initialdischarge pressure, a first discharge pressure, and a second dischargepressure are respectively denoted by P₀, P₁, and P₂, and errors betweenthe pressures are denoted by D1 and D2 respectively, equations P₁=P₀+D₁and P₂=P₁+D₂ hold. Accordingly, as shown in the equation of FIG. 14, ann-th discharge pressure includes an accumulated error corresponding to ntimes discharges.

FIG. 1 is a schematic of a microinjection apparatus 100 according to afirst embodiment of the present invention. FIGS. 2A to 2C are schematicsfor explaining an overview and features of a liquid discharge volumecontrol method according to the first embodiment.

The microinjection apparatus 100, which is connected to the positivepressure pump 1 and the negative pressure pump 2, includes the regulator3, the regulating chamber 5, the capillary 9, the valve a4, the valveb7, and the pressure sensor a6. The regulator 3 maintains a pressuredeveloped by the pumps 1 and 2 constant. The regulating chamber 5 trapsthe pressure maintained by the regulator 3. The capillary 9 is filledwith a solution. The valve a4 is between the regulator 3 and theregulating chamber 5. The valve b7 is between the regulating chamber 5and the capillary 9. The pressure sensor a6 detects a pressure in theregulating chamber 5. The regulator 3, the valve a4, the regulatingchamber 5, the pressure sensor a6, the valve b7, and the capillary 9 areconnected by the tube 10 (indicated by thick lines). The regulatingchamber 5 referred to here is a portion of the tube 10 that connectsbetween the valve a4 and the pressure sensor a6 and between the pressuresensor a6 and the valve b7. The liquid-discharge-volume control device20 controls operations of the microinjection apparatus 100. For example,as each of the valves a4 and b7, a valve including an electromagneticsolenoid is employed. The liquid-discharge-volume control device 20generates an electric signal, thereby controlling opening and closingoperations of each of the valves a4 and b7.

While the liquid-discharge-volume control device 20 of the firstembodiment is capable of controlling a liquid volume accurately as inthe conventional technology, the salient feature of theliquid-discharge-volume control device 20 is to provide more accurateand stable control of a liquid discharge volume.

The salient feature is briefly explained. First, theliquid-discharge-volume control device 20 of the first embodimentperforms motion control of the microinjection apparatus 100 as shown inFIG. 2A to apply the injection pressure Pi to the capillary 9 to startdischarge of liquid. Specifically, the liquid-discharge-volume controldevice 20 causes the regulator 3 to generate the maintaining pressure Pcby using the positive pressure pump 1 and the negative pressure pump 2,and then opens the valves a4 and b7 (see (1) of FIG. 2A). Next, anoperator connects the capillary 9 filled with a solution to the tube 10.Hence, the liquid filled in the capillary 9 is retained in the capillary9 without flowing backward. The state shown in (1) of FIG. 2A ismaintained until penetration of a cell with the capillary 9 by theoperator under a microscope to discharge (to inject) the liquid becomesready.

When penetration of the cell with the capillary 9 by the operator tostart injection becomes available, as shown in (2) of FIG. 2A, theliquid-discharge-volume control device 20 closes the valve b7 tomaintain the capillary 9 under the maintaining pressure Pc in accordancewith an operator request to start injection, and causes the regulator 3to generate the injection-pressure generating pressure Ph (describedlater), which is a pressure necessary to generate the injection pressurePi, by using the positive pressure pump 1 and the negative pressure pump2 (see (3) of FIG. 2A).

As shown in (4) of FIG. 2A, the liquid-discharge-volume control device20 closes the valve a4 to maintain the regulating chamber 5 under theinjection-pressure generating pressure Ph, and then opens the valve b7.Consequently, the injection-pressure generating pressure Ph and themaintaining pressure Pc are combined into a combined pressure Pi′, andliquid is discharged out of the capillary 9 (see (5) of FIG. 2A). Asexplained above, instantaneous opening/closing of the valves a4 and b7causes the pressure applied to the capillary to change from themaintaining pressure to the injection pressure quickly. The series ofoperations are also disclosed in Japanese Patent Application Laid-OpenNo. 2006-133512, “METHOD FOR DISCHARGING LIQUID INTO CELL ANDMICROINJECTION APPARATUS” by the inventor of this application.

However, a change in the “volume ratio η” caused by elements such asflexibility of the material of the tube 10, pressure values, and apressure-applying time can develop a slight error in the obtainedcombined pressure Pi′ in relation to the injection pressure Pi.

Therefore, as shown in (6) of FIG. 2B, the liquid-discharge-volumecontrol device 20 of the first embodiment sets an output of theregulator 3 to the injection pressure Pi, thereby reapplying theinjection pressure Pi to the capillary 9. This corrects the errorincluded in the combined pressure Pi′, which is obtained at step (5) ofFIG. 2A, in relation to the injection pressure Pi.

Subsequently, as shown in (7) of FIG. 2B, the liquid-discharge-volumecontrol device 20 of the first embodiment closes the valve b7 tomaintain the capillary 9 under the injection pressure Pi, and causes theregulator 3 to generate the maintaining-pressure generating pressure PL(described later), which is necessary to generate the maintainingpressure Pc (see (8) of FIG. 2B).

As shown in (9) of FIG. 2B, the liquid-discharge-volume control device20 closes the valve a4 to maintain the regulating chamber 5 under themaintaining-pressure generating pressure PL, and then opens the valve b7(see (10) of FIG. 2B). Consequently, the maintaining-pressure generatingpressure PL and the injection pressure Pi are combined into the combinedpressure Pc′, which terminates the discharge of liquid out of thecapillary 9. As explained above, instantaneous opening/closing of thevalves a4 and b7 causes the pressure applied to the capillary to changefrom the injection pressure to the maintaining pressure quickly.Accordingly, an injection pressure that takes a rectangular waveform canbe applied to the capillary 9, which allows to perform accurate controlof a liquid discharge volume by adjusting the injection pressure and aninjection-pressure application time (a period of time required toprocess steps (5) of FIG. 2A to (10) of FIG. 2B). Meanwhile, as in thecase of the combined pressure Pi′, the combined pressure Pc′ can have aslight error in relation to the maintaining pressure Pc.

When the liquid-discharge-volume control device 20 is required toconsecutively discharge liquid to another cell, theliquid-discharge-volume control device 20 causes the regulator 3 togenerate the maintaining pressure Pc, and opens the valve a4 to shiftthe state from that shown in (10) of FIG. 2B to that shown in (1) ofFIG. 2A, to perform operation control including the same steps as above.Shifting from step (10) of FIG. 2B to step (1) of FIG. 2A causes theprocess to enter standby mode until a start of a next injection cycle,and corrects the error, which can be included in the combined pressurePc′ obtained at step (10) of FIG. 2B, in relation to the maintainingpressure Pc. Therefore, this operation achieves reapplication of themaintaining pressure to the capillary 9 even when the process is notfollowed by another injection.

FIG. 2C is a graph that depicts open/close state of the valves a4 andb7, pressure output from the regulator 3, pressure applied to theregulating chamber 5, and pressure applied to the capillary 9 in (1) to(5) of FIG. 2A and (6) to (9) of FIG. 2B together. The numbers 1 to 10(STEP) on the horizontal axis correspond to steps (1) to (10) of FIGS.2A and 2B, respectively.

Therefore, even when an injection pressure or a maintaining pressureobtained as above involves an error, the liquid discharge volume controlmethod of the first embodiment can absorb the error by reapplying theinjection pressure or the maintaining pressure to the capillary 9.Hence, as previously explained as the salient feature, the methodachieves more accurate and stable control of a liquid discharge volume.

Meanwhile, in contrast to the microinjection apparatus 1000, themicroinjection apparatus 100 does not include the pressure sensor b8.This is because, since a pressure value detected by the pressure sensora6 with the valve b7 opened is identical with a pressure value (pressureon the capillary side) detected by the pressure sensor b8 at the samepoint in time, a pressure value detected by the pressure sensor a6immediately before the valve b7 is closed is employed as thecapillary-side pressure so that the pressure sensor a6 functions also asthe pressure sensor b8.

The above-mentioned injection-pressure generating pressure Ph forgenerating the injection pressure Pi and the maintaining-pressuregenerating pressure PL for generating the maintaining pressure Pc areexplained. A relation between the pressure at a point in time before thevalve b7 is opened and the pressure at a point in time after the valveb7 is opened can be obtained using an equation of state of gas. As shownin FIG. 1, the gas volume on the capillary 9 side is denoted as V1, thepressure on the capillary 9 side at a point in time before the valve b7is opened as P1, the gas volume of the regulating chamber 5 as V2, andthe pressure in the regulating chamber 5 at a point in time before thevalve b7 is opened as P2. In this case, relations between the pressureP, which is a pressure at a point in time after the valve b7 is opened,and the volume ratio η (η=V2/N1) are expressed in each of equations (1)and (2) of FIG. 13. Accordingly, the pressure P2 that need to beretained in the regulating chamber 5 before the valve b7 is opened togenerate the pressure P after the valve b7 is opened can be expressed byequation (3) of FIG. 13.

Meanwhile, the pressure P1 on the capillary 9 side between closing andopening of the valve b7 can be obtained as a pressure value detected bythe pressure sensor a6 immediately before the valve b7 is closed; thepressure P2 of the regulating chamber 5 at a point in time before thevalve b7 is opened can be obtained using the pressure sensor a6; and thepressure P at a point in time after the valve b7 is opened can beobtained using the pressure sensor a6. Therefore, the volume ratio η canbe obtained in advance from equation (2) of FIG. 13 using the pressurevalues determined by performing a preliminary discharge of liquid.

The injection-pressure generating pressure Ph or themaintaining-pressure generating pressure PL is calculated using thevolume ratio η and equation (3) of FIG. 13. The injection-pressuregenerating pressure Ph is calculated by substituting the injectionpressure Pi for the pressure P, which is the pressure at a point in timeafter the valve b7 is opened, and the maintaining pressure Pc for thepressure P1, which is the pressure on the capillary 9 side at a point intime before the valve b7 is opened, in equation (3) of FIG. 13. Themaintaining-pressure generating pressure PL is calculated bysubstituting the maintaining pressure Pc for the pressure P, which isthe pressure at a point in time after the valve b7 is opened, and theinjection pressure Pi for the pressure P1, which is the pressure on thecapillary 9 side at a point in time before the valve b7 is opened, inequation (3) of FIG. 13.

FIG. 3 is a block diagram of the liquid discharge volume control device20 according to the first embodiment. The liquid-discharge-volumecontrol device 20 includes an input unit 21, an output unit 22, aninput/output control interface (I/F) 23, a calculating unit 24, astorage unit 25, and a controller 26, and is connected to themicroinjection apparatus 100.

The input unit 21 includes a keyboard, a mouse, and the like, andreceives an operator input on various conditions (e.g., a liquiddischarge volume to be set based on the injection pressure Pi and apressure-application time, and the maintaining pressure Pc to be setbased on the viscosity of a liquid filled in the capillary 9, and thelike) pertaining to injection, and an operator request to startinjection.

The output unit 22 outputs a control instruction issued by thecontroller 26, described later, to the microinjection apparatus 100, andcontrols operations of the microinjection apparatus 100.

The input/output control I/F 23 controls data transfer to and from theinput unit 21, the output unit 22, the calculating unit 24, the storageunit 25, and the controller 26.

The storage unit 25 stores therein data to be used in various processesperformed by the calculating unit 24, results of the various processesperformed by the calculating unit 24, and records of various controloperations performed by the controller 26. The storage unit 25 includesa pressure sensor-output storage unit 251, a calculated-volume-ratiostorage unit 252, an injection-pressure-generating-pressure storage unit253, and a maintaining-pressure-generating-pressure storage unit 254.

The pressure sensor-output storage unit 251 stores therein, in additionto a pressure value obtained from the pressure sensor a6 through apressure sensor-output detector 261, described later, a record ofopening/closing control on the valve a4 performed by a valve a4controller 263, described later, and a record of opening/closing controlon the valve b7 performed by a valve b7 controller 264, described later.

The calculated-volume-ratio storage unit 252 stores therein a volumeratio calculated by a volume-ratio calculating unit 241, describedlater. The injection-pressure-generating-pressure storage unit 253stores therein an injection-pressure generating pressure calculated byan injection-pressure-generating-pressure calculating unit 242,described later. The maintaining-pressure-generating-pressure storageunit 254 stores therein a maintaining-pressure generating pressurecalculated by a maintaining-pressure-generating-pressure calculatingunit 243, described later. The respective units are described in detaillater.

The controller 26 performs various control operations in accordance withan operator request transmitted from the input/output control I/F 23.The controller 26 includes the pressure sensor-output detector 261, aregulator-output-pressure controller 262, the valve a4 controller 263,and the valve b7 controller 264.

The pressure sensor-output detector 261 obtains a pressure valuedetected by the pressure sensor a6 from the pressure sensor a6, andstores therein the result in the pressure sensor-output storage unit251.

The regulator-output-pressure controller 262 causes the regulator 3 togenerate a pressure of a predetermined magnitude by using the positivepressure pump 1 and the negative pressure pump 2. Put another way, theregulator-output-pressure controller 262 causes the regulator 3 togenerate the injection pressure Pi or the maintaining pressure Pc, whichis a setting condition input to the input unit 21, theinjection-pressure generating pressure Ph stored in theinjection-pressure-generating-pressure storage unit 253, or themaintaining-pressure generating pressure PL stored in themaintaining-pressure-generating-pressure storage unit 254.

The valve a4 controller 263 controls opening and closing operations ofthe valve a4. Specifically, the valve a4 controller 263 controls theseries of opening and closing operations of the valve a4 shown in FIGS.2A and 2B by using, e.g., electric signals. The valve a4 controller 263also transmits a record of opening/closing control on the valve a4 tothe pressure sensor-output storage unit 251. The pressure sensor-outputstorage unit 251 stores therein the record of opening/closing control onthe valve a4.

The valve b7 controller 264 controls opening closing operations of thevalve b7. The valve b7 controller 264 also transmits a record ofopening/closing control on the valve b7 to the pressure sensor-outputstorage unit 251. The pressure sensor-output storage unit 251 storestherein the record of opening/closing control on the valve b7.

The calculating unit 24 performs various processes in accordance with anoperator request transmitted from the input/output control I/F 23. Thecalculating unit 24 includes the volume-ratio calculating unit 241, theinjection-pressure-generating-pressure calculating unit 242, and themaintaining-pressure-generating-pressure calculating unit 243.

The volume-ratio calculating unit 241 calculates the volume ration froma record on an output pressure value, which is detected by the pressuresensor a6 and stored in the pressure sensor-output storage unit 251, acontrol record of the valve a4 controller 263, and a control record ofthe valve b7 controller 264, and stores the calculation result in thecalculated-volume-ratio storage unit 252.

Calculation of a volume ratio is explained in detail. The valve a4controller 263 opens the valve a4, the valve b7 controller 264 opens thevalve b7, and the regulator-output-pressure controller 262 causes theregulator 3 to generate an appropriate pressure P1. The valve b7controller 264 then closes the valve b7 to maintain the capillary 9under the pressure P1. As the value of the pressure P1, a pressure valuedetected by the pressure sensor a6 immediately before the valve b7 isclosed is employed by referring to the control record of the valve b7controller 264. The regulator-output-pressure controller 262 causes theregulator 3 to generate an appropriate pressure P2, and the valve a4controller 263 closes the valve a4 to maintain the regulating chamber 5under the pressure P2. As the value of the pressure P2, a value detectedby the pressure sensor a6 at this stage is employed by referring to thecontrol record of the valve a4 controller 263 and the control record ofthe valve b7 controller 264. Subsequently, the valve b7 controller 264opens the valve b7 to combine the pressure P1 with the pressure P2 togenerate a new pressure P. The pressure P is applied to the capillary 9.As the value of the pressure P, a value detected by the pressure sensora6 at this stage is employed by referring to the control record of thevalve a4 controller 263 and the control record of the valve b7controller 264. The volume ratio η is obtained from the actual values ofP1, P2, and P by using equation (2) of FIG. 13.

The injection-pressure-generating-pressure calculating unit 242calculates the injection-pressure generating pressure Ph, which isnecessary to obtain the injection pressure Pi in combination with themaintaining pressure Pc, by using the volume ratio η stored in thecalculated-volume-ratio storage unit 252, and stores the calculationresult in the injection-pressure-generating-pressure storage unit 253.Specifically, injection-pressure-generating-pressure calculating unit242 uses the volume ratio η to calculate the injection-pressuregenerating pressure Ph, and substitutes the injection pressure Pi forthe pressure P, which is the pressure at a point in time after the valveb7 is opened, and the maintaining pressure Pc for the pressure P1, whichis the pressure on the capillary 9 side at a point in time before thevalve b7 is opened, in equation (3) of FIG. 13. Thus, the followingequation is formed:

Ph=((η+1)Pi−Pc)/η.

The maintaining-pressure-generating-pressure calculating unit 243calculates the maintaining-pressure generating pressure PL, which isnecessary to generate the maintaining pressure Pc in combination withthe injection pressure Pi, by using the volume ratio η stored in thecalculated-volume-ratio storage unit 252, and stores the calculationresult in the maintaining-pressure-generating-pressure storage unit 254.Specifically, the maintaining-pressure-generating-pressure calculatingunit 243 calculates the maintaining-pressure generating pressure PL bysubstituting the maintaining pressure Pc for the pressure P, which isthe pressure at a point in time after the valve b7 is opened, and theinjection pressure Pi for the pressure P1, which is the pressure on thecapillary 9 side at a point in time before the valve b7 is opened, inequation (3) of FIG. 13. Thus, the following equation is formed:

Ph=((η+1)Pc−Pi)/η.

FIG. 4 is a flowchart of a process procedure performed by the liquiddischarge volume control device 20.

First, when the liquid-discharge-volume control device 20 receives a newoperator input on various conditions (e.g., a liquid discharge volume tobe set based on the injection pressure Pi and a pressure-applicationtime, and the maintaining pressure Pc to be set based on the viscosityof a liquid filled in the capillary 9, and the like) pertaining toinjection via the keyboard or the mouse (YES at step S401), theinjection-pressure-generating-pressure calculating unit 242 and themaintaining-pressure-generating-pressure calculating unit 243 calculatethe injection-pressure generating pressure Ph and themaintaining-pressure generating pressure PL, respectively (step S402).

While the regulator-output-pressure controller 262 causes the regulator3 to generate the maintaining pressure Pc by using the positive pressurepump 1 and the negative pressure pump 2, the valve a4 controller 263opens the valve a4, and the valve b7 controller 264 opens the valve b7(step S403). Hence, the liquid filled in the capillary 9 is preventedfrom flowing backward into the capillary 9.

Thereafter, the liquid-discharge-volume control device 20 maintains thestate until when penetration of a cell with the capillary 9 by theoperator to start injection becomes available and theliquid-discharge-volume control device 20 receives an operator requestto start injection (NO at step S404).

On the other hand, when penetration of a cell with the capillary 9 bythe operator to start injection becomes available and theliquid-discharge-volume control device 20 receives an operator requestto start injection (YES at step S404), the valve b7 controller 264closes the valves b7, and the regulator-output-pressure controller 262causes the regulator 3 to generate the injection-pressure generatingpressure Ph by using the positive pressure pump 1 and the negativepressure pump 2 (step S405).

While the valve a4 controller 263 closes the valve a4 to maintain theregulating chamber 5 under the injection-pressure generating pressurePh, the valve b7 controller 264 opens the valve b7 to combine theinjection-pressure generating pressure Ph with the maintaining pressurePc. The newly-generated combined pressure is applied to the capillary 9(step S406). Consequently, the pressure applied to the capillary 9increases quickly to start discharging of liquid out of the capillary 9.

Subsequently, while the regulator-output-pressure controller 262 causesthe regulator 3 to generate the injection pressure Pi by using thepositive pressure pump 1 and the negative pressure pump 2, the valve a4controller 263 opens the valve a4 (step S407). This operation allows,even when the combined pressure obtained at step S406 involves an errorin relation to the injection pressure Pi, to correct the error, and toapply the injection pressure Pi to the capillary 9.

While the valve b7 controller 264 closes the valves b7 (step S408),thereby maintaining the capillary 9 under the injection pressure Pi, theregulator-output-pressure controller 262 causes the regulator 3 togenerate the maintaining-pressure generating pressure PL by using thepositive pressure pump 1 and the negative pressure pump 2 (step S409).

While the valve a4 controller 263 closes the valve a4 to maintain theregulating chamber 5 under the maintaining-pressure generating pressurePL, the valve b7 controller 264 opens the valve b7 to combine themaintaining-pressure generating pressure PL with the injection pressurePi. The newly-generated combined pressure is applied to the capillary 9(step S410). Consequently, the pressure applied to the capillary 9decreases quickly to terminate the discharge of liquid out of thecapillary 9.

While the regulator-output-pressure controller 262 causes the regulator3 to generate the maintaining pressure Pc by using the positive pressurepump 1 and the negative pressure pump 2, the valve a4 controller 263opens the valve a4 (step S411). This operation allows, even when thecombined pressure obtained at step S410 involves an error in relation tothe maintaining pressure Pc, to correct the error, and to apply themaintaining pressure Pc to the capillary 9.

Upon receipt of a new operator request to start injection (YES at stepS412), the liquid-discharge-volume control device 20 performs theprocess pertaining to step S405 and subsequent steps. When a new requestfor injection start is not issued (NO at step S412), the process ends.

As explained above, according to the first embodiment, opening the valveb7 in the state where the valves a4 and b7 are closed causes aninjection-pressure generating pressure trapped between the valves a4 andb7 and a maintaining pressure applied to the capillary 9 to be combinedinto an injection pressure. Hence, discharge of the liquid out of thecapillary 9 is started by the injection pressure. Thereafter, while anoutput pressure of the regulator 3 is set to the injection pressure, thevalve a4 is opened to apply the injection pressure to the capillary 9,and then the valve b7 is opened in a state where the valves a4 and b7are closed. Accordingly, a maintaining-pressure generating pressuretrapped between the valves a4 and b7 and the injection pressure appliedto the capillary 9 are combined into a maintaining pressure. Hence, thedischarge of liquid is terminated by the maintaining pressure.Thereafter, while the output pressure of the regulator 3 is set to themaintaining pressure, the valve a4 is opened to apply the maintainingpressure to the capillary 9. Therefore, even when, for example, theinjection pressure or the maintaining pressure generated as aboveinvolves an error, the error can be absorbed by reapplying the injectionpressure or the maintaining pressure to the capillary 9, therebyachieving more accurate and stable control of a liquid discharge volume.

According to the first embodiment, calculation of a volume ratio isperformed by employing, as an actual value of the pressure applied tothe capillary 9, a value detected by the pressure sensor a6 immediatelybefore the valve b7 is closed, and employing, as a value of the combinedpressure generated when the valve b7 is opened, a value detected by thepressure sensor a6. Therefore, the magnitude of the pressure output tothe capillary can be detected without provision of another pressuresensor for detecting the pressure, thereby suppressing a cost of capitalinvestment.

In the first embodiment, the injection-pressure generating pressure Phor the maintaining-pressure generating pressure PL is calculated basedon a previously-calculated volume ratio. In a second embodiment of thepresent invention, a volume ratio to be borne under a pressure conditiondefined by input is recalculated to calculate the injection-pressuregenerating pressure Ph or the maintaining-pressure generating pressurePL.

FIG. 5 is a schematic for explaining an overview and features of theliquid discharge volume control device of the second embodiment.

As in the first embodiment, the liquid discharge volume control deviceof the second embodiment calculates the injection-pressure generatingpressure Ph and the maintaining-pressure generating pressure PL from theinjection pressure Pi and the maintaining pressure Pc, which are inputby an operator, using the already-calculated volume ratio η and equation(3) of FIG. 13.

The liquid discharge volume control device of the second embodimentcontrols operations of the microinjection apparatus 100 to open thevalve b7 in a state where the injection-pressure generating pressure Phis maintained in the regulating chamber 5 and the capillary 9 receivesthe maintaining pressure Pc as shown in the upper diagram in (A) of FIG.5, thereby generating a combined pressure as shown in the lower diagramin (A) of FIG. 5. At this timing, an actual value of theinjection-pressure generating pressure Ph, that of the maintainingpressure Pc, and a value of the combined pressure Pi′ are obtained usingthe pressure sensor a6. As the actual value of the maintaining pressurePc, a value detected by the pressure sensor a6 immediately beforeclosing of the valve b7 causes the state to shift to that shown in theupper diagram in (A) of FIG. 5 is employed.

Subsequently, the liquid discharge volume control device of the secondembodiment calculates an injection-pressure-compensating volume ratioηi, which is a volume ratio to be borne under a condition where the tube10 receives the maintaining pressure Pc and the injection-pressuregenerating pressure Ph, by using equation (2) of FIG. 13. Specifically,the actual value Pi′ is substituted for P, an actual value of themaintaining pressure Pc is substituted for P1, and an actual value ofthe injection-pressure generating pressure Ph is substituted for P2 inequation (2) of FIG. 13. Thus, the following equation is formed:

η=((Pc−Pi′)/(P′−Ph)

where Pc and Ph are actual values. A compensated injection-pressuregenerating pressure Ph′, which is necessary to generate a renewedinjection pressure Pi, is calculated using theinjection-pressure-compensating volume ratio ηi. Specifically, withequation (3) of FIG. 13, the compensated injection-pressure generatingpressure Ph′ is calculated by substituting Pi, which is a pressure valueof an operator input, for P, the maintaining pressure Pc for P1, and theinjection-pressure-compensating volume ratio ηi for the volume ratio η,i.e., Ph′=((ηi+1)(Pi−Pc)/ηi.

The liquid discharge volume control device of the second embodimentcontrols operations of the microinjection apparatus 100 to open thevalve b7 in a state where the maintaining-pressure generating pressurePL is maintained in the regulating chamber 5 and the capillary 9receives the injection pressure Pi as shown in the upper diagram in (B)of FIG. 5, thereby obtaining a combined pressure as shown in the lowerdiagram in (B) of FIG. 5. At this timing, an actual value of themaintaining-pressure generating pressure PL, that of the injectionpressure Pi, and a value of the combined pressure Pi′ are obtained usingthe pressure sensor a6. As the actual value of the injection pressurePi, a value detected by the pressure sensor a6 immediately beforeclosing of the valve b7 causes the state to shift to that shown in theupper diagram in (B) of FIG. 5 is employed.

Subsequently, the liquid discharge volume control device of the secondembodiment calculates a maintaining-pressure-compensating volume ratioηc, which is a volume ratio to be borne under a condition where the tube10 receives the injection pressure Pi and the maintaining-pressuregenerating pressure PL, by using equation (2) of FIG. 13. Specifically,the actual value Pc′ is substituted for P, an actual value of theinjection pressure Pi is substituted for P1, and an actual value of themaintaining-pressure generating pressure PL is substituted for P2 inequation (2) of FIG. 13. Thus, the following equation is formed:

η=((Pi−Pc′)/(Pc′−PL)

where Pi and PL are actual values.

A compensated maintaining-pressure generating pressure PL′, which isnecessary to generate a renewed maintaining pressure Pc, is calculatedusing the maintaining-pressure-compensating volume ratio ηc.Specifically, with equation (3) of FIG. 13, the compensatedmaintaining-pressure generating pressure PL′ is calculated bysubstituting Pc, which is a pressure value of an operator input, for P,the injection pressure Pi for P1, and themaintaining-pressure-compensating volume ratio ηc for the volume ratioη, i.e., PL′=((ηc+1)Pc−Pi)/η.

The liquid discharge volume control device of the second embodimentperforms discharge of liquid into a cell by using the compensatedinjection-pressure generating pressure Ph′ and the compensatedmaintaining-pressure generating pressure PL′ in accordance with anoperator request to start injection.

In this embodiment, the compensated injection-pressure generatingpressure Ph′ and the compensated maintaining-pressure generatingpressure PL′ are calculated based on an actual combined pressure value,and the like, obtained through a single series of operations. However,alternatively, the compensated injection-pressure generating pressurePh′ and the compensated maintaining-pressure generating pressure PL′ canbe calculated based on an average combined pressure value obtained byrepeating the series of operations shown in (A) and (B) of FIG. 5 aplurality of times.

As explained above, the liquid discharge volume control device of thesecond embodiment is capable of recalculating a magnitude of pressure tobe output from the regulator to generate an injection pressure or amaintaining pressure while compensating a volume ratio based on anactual pressure value taken for each injection setting condition.Accordingly, stabilization of liquid discharge volume is achieved underany setting condition.

Next, the liquid discharge volume control device according to the secondembodiment is explained with reference to FIG. 3. The liquid dischargevolume control device 20 according to the second embodiment is basicallysimilar to that of the first embodiment except that the volume-ratiocalculating unit 241, the injection-pressure-generating-pressurecalculating unit 242, and the maintaining-pressure-generating-pressurecalculating unit 243 operate differently from those in the firstembodiment. Therefore, the same explanation is not repeated.

The volume-ratio calculating unit 241 recalculates theinjection-pressure-compensating volume ratio ηi and theinjection-pressure-compensating volume ratio ηc, and stores thecalculation results in the calculated-volume-ratio storage unit 252.That is, the volume-ratio calculating unit 241 calculates theinjection-pressure-compensating volume ratio ηi, which is a volume ratioto be borne under a condition where the tube 10 receives the maintainingpressure Pc and the injection-pressure generating pressure Ph, based onthe actual pressure value detected in the step shown in (A) of FIG. 5and stored in the pressure sensor-output storage unit 251 by usingequation (2) of FIG. 13, and stores the calculation result in thecalculated-volume-ratio storage unit 252. Specifically, theinjection-pressure-compensating volume ratio ηi is calculated bysubstituting the actual value Pi′ for P, an actual value of themaintaining pressure Pc for P1, and an actual value of theinjection-pressure generating pressure Ph for P2 in equation (2) of FIG.13.

The volume-ratio calculating unit 241 calculates themaintaining-pressure-compensating volume ratio ηc, which is a volumeratio to be borne under a condition where the tube 10 receives themaintaining-pressure generating pressure PL and the injection pressurePi, based on the actual pressure value detected in the step shown in (B)of FIG. 5 and stored in the pressure sensor-output storage unit 251 byusing equation (2) of FIG. 13, and stores the calculation result in thecalculated-volume-ratio storage unit 252. Specifically, themaintaining-pressure-compensating volume ratio ηc is calculated bysubstituting the actual value Pi′ for P, an actual value of theinjection pressure Pc for P1, and an actual value of themaintaining-pressure generating pressure PL for P2 in equation (2) ofFIG. 13.

The injection-pressure-generating-pressure calculating unit 242calculates the compensated injection-pressure generating pressure Ph′,which is necessary to generate a renewed injection pressure Pi, usingthe injection-pressure-compensating volume ratio ηi stored in thecalculated-volume-ratio storage unit 252. Specifically, with equation(3) of FIG. 13, the compensated injection-pressure generating pressurePh′ is calculated by substituting Pi, which is a pressure value of anoperator input, for P, the maintaining pressure Pc for P1, and theinjection-pressure-compensating volume ratio ηi for the volume ratio η.

The maintaining-pressure-generating-pressure calculating unit 243calculates the compensated maintaining-pressure generating pressure PL′,which is necessary to generate a renewed maintaining pressure Pc, usingthe maintaining-pressure-compensating volume ratio ηc stored in thecalculated-volume-ratio storage unit 252. Specifically, with equation(3) of FIG. 13, the compensated maintaining-pressure generating pressurePL′ is calculated by substituting Pc, which is a pressure value of anoperator input, for P, the injection pressure Pi for P1, and themaintaining-pressure-compensating volume ratio ηc for the volume ratioη.

FIG. 6 is a flowchart of a process procedure performed by the liquiddischarge volume control device according to the second embodiment.

First, when the liquid-discharge-volume control device 20 of the secondembodiment receives a new operator input on various conditions (e.g., aliquid discharge volume to be set based on the injection pressure Pi anda pressure-application time, and the maintaining pressure Pc to be setbased on the viscosity of a liquid filled in the capillary 9, and thelike) pertaining to injection via a keyboard or a mouse (YES at stepS601), the same process at steps S402 to S403 in FIG. 3 as describedpreviously in the first embodiment is performed (steps S602 to S603).

Subsequently, the liquid-discharge-volume control device 20 of thesecond embodiment maintains the state until an operator request tocalculate a compensated injection-pressure generating pressure and acompensated maintaining-pressure generating pressure-compensatingpressure is received (NO at step S604).

On the other hand, when the liquid-discharge-volume control device 20 ofthe second embodiment receives an operator calculation request tocalculate a compensated injection-pressure generating pressure and acompensated maintaining-pressure generating pressure-compensatingpressure (YES at step S604), the valve b7 controller 264 closes thevalves b7, and the regulator-output-pressure controller 262 causes theregulator 3 to generate the injection-pressure generating pressure Ph byusing the positive pressure pump 1 and the negative pressure pump 2(step S605).

While the valve a4 controller 263 closes the valve a4 to maintain theregulating chamber 5 under the injection-pressure generating pressurePh, the valve b7 controller 264 opens the valve b7 to combine theinjection-pressure generating pressure Ph with the maintaining pressurePc. The newly-generated combined pressure is applied to the capillary 9(step S606).

The pressure sensor-output detector 261 obtains the actual value Pi′ ofthe newly-generated combined pressure, and the like, and stores thevalues in the pressure sensor-output storage unit 251 (step S607). Thevolume-ratio calculating unit 241 calculates theinjection-pressure-compensating volume ratio ηi using the actual valuestored in the pressure sensor-output storage unit 251, and theinjection-pressure-generating-pressure calculating unit 242 calculatesthe compensated injection-pressure generating pressure Ph′ using theinjection-pressure-compensating volume ratio ηi (step S608).

Subsequently, while the regulator-output-pressure controller 262 causesthe regulator 3 to generate the injection pressure Pi by using thepositive pressure pump 1 and the negative pressure pump 2, the valve a4controller 263 opens the valve a4 (step S609).

While the valve b7 controller 264 closes the valves b7 (step S610) tomaintain the capillary 9 under the injection pressure Pi, theregulator-output-pressure controller 262 causes the regulator 3 togenerate the maintaining-pressure generating pressure PL by using thepositive pressure pump 1 and the negative pressure pump 2 (step S611).

While the valve a4 controller 263 closes the valve a4 to maintain theregulating chamber 5 under the maintaining-pressure generating pressurePL, the valve b7 controller 264 opens the valve b7 to combine themaintaining-pressure generating pressure PL with the injection pressurePi. The newly-generated combined pressure is applied to the capillary 9(step S612).

The pressure sensor-output detector 261 obtains an actual value Pc′ ofthe newly-generated combined pressure, and the like, and stores thevalues in the pressure sensor-output storage unit 251 (step S613). Thevolume-ratio calculating unit 241 calculates themaintaining-pressure-compensating volume ratio ηc using the actual valuestored in the pressure sensor-output storage unit 251, and themaintaining-pressure-generating-pressure calculating unit 243 calculatesthe compensated maintaining-pressure generating pressure PL′ using themaintaining-pressure-compensating volume ratio ηc (step S614).

While the regulator-output-pressure controller 262 causes the regulator3 to generate the maintaining pressure Pc by using the positive pressurepump 1 and the negative pressure pump 2, the valve a4 controller 263opens the valve a4 (step S615).

Upon receipt of an operator request to start injection (YES at stepS616), the liquid-discharge-volume control device 20 of the secondembodiment performs the process pertaining to step S405 and subsequentsteps. When a new request for injection start is not issued (NO at stepS616), the process ends.

As explained above, according to the second embodiment, aninjection-pressure generating pressure is compensated by detecting anactual value of pressure output from the regulator 3 as theinjection-pressure generating pressure with the pressure sensor a6, anactual value of pressure applied to the capillary 9 as a maintainingpressure with the pressure sensor a6 immediately before the valve b7 isclosed, and a value of a combined pressure generated by opening thevalve b7 with the pressure sensor a6; and a maintaining-pressuregenerating pressure is compensated by detecting an actual value ofpressure output from the regulator 3 as the maintaining-pressuregenerating pressure with the pressure sensor a6, an actual value ofpressure applied to the capillary 9 as an injection pressure with thepressure sensor a6 immediately before the valve b7 is closed, and avalue of a generated combined pressure with the pressure sensor a6.Therefore, it is possible to recalculate a value of pressure to beoutput from the regulator 3 to generate a renewed injection pressure ora renewed maintaining pressure based on the actual pressure values takenfor each injection setting condition, thereby achieving stabilization ofa liquid discharge volume under any setting condition.

FIG. 7 is a schematic for explaining an overview and features of aliquid-discharge-volume control device 30 according to a thirdembodiment of the present invention. The liquid-discharge-volume controldevice 30 is basically similar to the liquid discharge volume controldevice 20 except for its function of detecting air leakage. Therefore,the same explanation is not repeated.

As with the liquid discharge volume control device 20, theliquid-discharge-volume control device 30 calculates theinjection-pressure generating pressure Ph and the maintaining-pressuregenerating pressure PL based on the injection pressure Pi and themaintaining pressure Pc, which are input by an operator, using thealready-calculated volume ratio η and equation (3) of FIG. 13. Theliquid-discharge-volume control device 20 causes the regulator 3 togenerate the maintaining pressure Pc by using the positive pressure pump1 and the negative pressure pump 2, and then opens the valves a4 and b7(see (A) of FIG. 7).

Subsequently, as shown in (B) of FIG. 7, the liquid discharge volumecontrol device closes the valve a4, acquires a value of pressure appliedto the capillary 9 after a predetermined period of time (e.g., after oneminute) from the pressure sensor a6, and subtracts the pressure valuefrom the maintaining pressure Pc to obtain a decrement. When thedecrement is smaller than a set threshold value, the liquid dischargevolume control device determines that no air leakage has occurred. Theliquid-discharge-volume control device 20 causes the regulator 3 togenerate the maintaining pressure Pc by using the positive pressure pump1 and the negative pressure pump 2, and then opens the valves a4 and b7to return to the state shown in (A) of FIG. 7, where theliquid-discharge-volume control device 20 waits while maintaining thestate until receipt of a request to start injection.

On the other hand, when the calculated decrement is equal to or largerthan the set threshold value, the liquid discharge volume control devicedetermines that air leakage has occurred, and transmits an alarm of thiseffect to the microinjection apparatus 100. This alarm allows anoperator to avoid starting injection in a state where air leakage hasoccurred due to poor connection of the capillary 9 or deterioration ofthe tube 10.

As explained above, because the liquid discharge volume control deviceof the third embodiment is capable of detecting air leakage due to poorconnection of a capillary or deterioration of a tube during preparationprior to injection, it is possible to avoid performing injection in astate where a liquid discharge volume is unstable due to the airleakage.

FIG. 8 is a block diagram of the liquid-discharge-volume control device30. The liquid-discharge-volume control device 30 includes a calculatingunit 240, a storage unit 250, and a controller 260 in place of thecalculating unit 24, the storage unit 25, and the controller 26. Thecalculating unit 240 further includes a maintaining-pressure-decrementcalculating unit 244. The storage unit 250 further includes amaintaining-pressure-decrement calculation-result storage unit 259, anda maintaining-pressure-decrement threshold-value storage unit 2510. Thecontroller 260 further includes an alarm transmitter 265.

As shown in (B) of FIG. 7, the maintaining-pressure-decrementcalculating unit 244 retrieves a value of pressure applied to thecapillary 9 from the pressure sensor-output storage unit 251 after alapse of a predetermined time (e.g., after one minute) since the valvea4 is closed. The maintaining-pressure-decrement calculating unit 244subtracts the pressure value from the maintaining pressure Pc to obtaina decrement, and stores the calculation result in themaintaining-pressure-decrement calculation-result storage unit 259.

The maintaining-pressure-decrement threshold-value storage unit 2510stores therein a set threshold value to be used in a process performedby the alarm transmitter 265, described later. Specifically, themaintaining-pressure-decrement threshold-value storage unit 2510 storestherein a set threshold value for use in determination as to whether airleakage has occurred.

The alarm transmitter 265 compares the decrement stored in themaintaining-pressure-decrement calculation-result storage unit 259 andthe set threshold value stored in the maintaining-pressure-decrementthreshold-value storage unit 2510. When the decrement is equal to orlarger than the set threshold value, the alarm transmitter 265determines that air leakage has occurred, and transmits an alarm of thiseffect to the microinjection apparatus 100. The alarm allows an operatorto avoid starting injection in a state where air leakage has occurreddue to poor connection of the capillary 9 or deterioration of the tube10.

FIG. 9 is a flowchart of a process procedure performed by theliquid-discharge-volume control device 30.

First, when the liquid-discharge-volume control device 30 receives a newoperator input on various conditions (e.g., a liquid discharge volume tobe set based on the injection pressure Pi and a pressure-applicationtime, and the maintaining pressure Pc to be set based on the viscosityof a liquid filled in the capillary 9, and the like) pertaining toinjection via a keyboard or a mouse (YES at step S901), the same processat steps S402 to S403 in FIG. 3 as described in the first embodiment isperformed (steps S902 to S903).

Subsequently, the valve a4 controller 263 closes the valve a4, retrievesa value of pressure applied to the capillary 9 from the pressuresensor-output storage unit 251 after a lapse of a predetermined time(e.g., after one minute) since the valve a4 is closed, and subtracts thepressure value from the maintaining pressure Pc to obtain a decrement(step S904).

The alarm transmitter 265 compares the decrement and the set thresholdvalue, thereby determining whether the decrement is equal to or largerthan the set threshold value (step S905). When the decrement is smallerthan the set threshold value (NO at step S905), while theregulator-output-pressure controller 262 causes the regulator 3 togenerate the maintaining pressure Pc by using the positive pressure pump1 and the negative pressure pump 2, the valve a4 controller 263 opensthe valve a4 (step S907) to maintain a state where the capillary 9receives the maintaining pressure Pc. The liquid-discharge-volumecontrol device 30 waits while maintaining this state until receipt of arequest to start injection (see step S404 shown in FIG. 4).

On the other hand, when the calculated decrement is equal to or largerthan the set threshold value (YES at step S905), the alarm transmitter265 determines that air leakage has occurred, and transmits an alarm ofthis effect to the microinjection apparatus 100 (step S906), and theprocess ends.

As explained above, according to the third embodiment, the valve a4 isclosed while the maintaining pressure generated by the regulator 3 uponopening of the valves a4 and b7 is being applied to the capillary 9.Then, the pressure sensor a6 measures a change in the maintainingpressure during a predetermined time period after the valve a4 is closedto detect pressure leakage. Therefore, air leakage due to poorconnection of the capillary 9 or deterioration of the tube 10 can bedetected during preparation prior to injection. Thus, it is possible toavoid performing injection in a state where a liquid discharge volume isunstable due to air leakage.

In the first to third embodiments, the microinjection apparatus 100includes the tube 10 of which volume changes depending on the magnitudeof pressure applied thereto. However, the invention is not so limited,and the tube 10 in the microinjection apparatus 100 can be formed of amaterial that exhibits no volume change. In this case, volumetric changeof the tube 10 that connects the regulating chamber 5, the valves a4 andb7, the capillary 9, and the like, can be suppressed, thereby achievingstabilization of a liquid discharge volume.

In the first and third embodiments, the injection-pressure generatingpressure Ph and the maintaining-pressure generating pressure PL arecalculated from the injection pressure Pi and the maintaining pressurePc, which are input by an operator, based on a volume ratio having beencalculated and stored. However, the present invention is not so limited,and, for example, it is possible to use a table that contains values ofthe injection-pressure generating pressure Ph corresponding to variousinjection pressures Pi obtained from calculation based on a volumeratio, and values of the maintaining-pressure generating pressure PLcorresponding to various maintaining pressures Pc obtained fromcalculation based on a volume ratio. An operator selects an injectionpressure and a maintaining pressure from a list of the table. Theliquid-discharge-volume control device refers to the table, andassociates the selected injection pressure and the maintaining pressureto an injection-pressure generating pressure and a maintaining-pressuregenerating pressure, respectively.

While, in the above embodiments, the microinjection apparatus iscontrolled, the present invention is so not limited, and can be appliedto any liquid dispensing apparatus that consecutively dispenses a givenvolume of liquid as well.

While, in the above embodiments, “solution” is discharged into a “cell”,the present invention is not so limited, and can be applied to the casethat “gas” is discharged into a “microstructure”.

Of the processes described in the embodiments, all or part of theprocesses explained as being performed automatically can be performedmanually (for example, occurrence of air leakage can be determined whenan operator inputs a request to start injection, instead of determiningthe air leakage after a lapse of a predetermined time). Similarly, allor part of the processes explained as being performed manually can beperformed automatically by a known method. The processing procedures,specific names, various data, and information including parametersdescribed in the embodiments or shown in the drawings can be changed asrequired unless otherwise specified. For example, the calculation of thecompensated injection-pressure generating pressure (step S608) can bestarted at the same time of calculating the compensatedmaintaining-pressure generating pressure (step S613).

The constituent elements of each device shown in the drawings arefunctionally conceptual, and need not be physically configured asillustrated. In other words, the specific mode (for example, the mode ofFIG. 3) of separation and integration of the constituent elements is notlimited to those shown in the drawings. The constituent elements, as awhole or in part, can be separated or integrated either functionally orphysically based on various types of loads or use conditions, forexample, the injection-pressure-generating-pressure calculating unit 242can be integrated with the maintaining-pressure-generating-pressurecalculating unit 243. The process functions performed by each device areentirely or partially realized by a central processing unit (CPU) orcomputer programs that are analyzed and executed by the CPU, or realizedas wired-logic hardware.

The liquid-discharge-volume control device is explained above ashardware; however, it can be implemented as software. In other words, acomputer program (hereinafter, a liquid discharge volume controlprogram) can be executed on a computer to realize the same function asthe liquid-discharge-volume control device. In the following, such acomputer that execute the liquid discharge volume control program torealize the same function as the liquid-discharge-volume control device20 in the first embodiment is explained. FIG. 10 is a diagram of acomputer 110 that executes the liquid discharge volume control program.

The computer 110 includes a keyboard 111, a display 112, a CPU 113, aread only memory (ROM) 114, a hard disk drive (HDD) 115, and a randomaccess memory (RAM) 116 and the like, which are connected by a bus 117.The computer 110 is further connected to the microinjection apparatus100.

In the ROM 114, the liquid discharge volume control program that fulfilsthe same function as the liquid-discharge-volume control device 20explained in the first embodiment. That is, as shown in FIG. 10, apressure sensor-output detection program 114 a, aregulator-output-pressure control program 114 b, a valve a4 controlprogram 114 c, a valve b7 control program 114 d, a volume-ratiocalculating program 114 e, an injection-pressure generating pressurecalculating program 114 f, and a maintaining-pressure generatingpressure calculating program 114 g, are stored in advance. The programs114 a to 114 g can be integrated or distributed as required.

The CPU 113 reads the programs 114 a to 114 g from the ROM 114 andexecutes the programs. Hence, the programs 114 a to 114 g function as apressure sensor-output detection process 113 a, aregulator-output-pressure control process 113 b, a valve a4 controlprocess 113 c, a valve b7 control process 113 d, a volume-ratiocalculating process 113 e, an injection-pressure generating pressurecalculating process 113 f, and a maintaining-pressure generatingpressure calculating process 113 g. The processes 113 a to 113 gcorrespond to the pressure sensor-output detector 261, theregulator-output-pressure controller 262, the valve a4 controller 263,the valve b7 controller 264, the volume-ratio calculating unit 241, theinjection-pressure-generating-pressure calculating unit 242, and themaintaining-pressure-generating-pressure calculating unit 243 shown inFIG. 2.

The HDD 115 includes pressure sensor-output data 115 a,calculated-volume-ratio data 115 b, injection-pressure generatingpressure data 115 c, and maintaining-pressure generating pressure data115 d. The pressure sensor-output data 115 a corresponds to the pressuresensor-output storage unit 251 explained with reference to FIG. 3, thecalculated-volume-ratio data 115 b corresponds to thecalculated-volume-ratio storage unit 252, the injection-pressuregenerating pressure data 115 c corresponds to the injection-pressuregenerating pressure storage unit 253, and the maintaining-pressuregenerating pressure data 115 d corresponds to the maintaining-pressuregenerating pressure storage unit 254. The CPU 113 stores the pressuresensor-output data 115 a as pressure sensor-output data 116 a, thecalculated-volume-ratio data 115 b as calculated-volume-ratio data 116b, the injection-pressure generating pressure data 115 c asinjection-pressure generating pressure data 116 c, and themaintaining-pressure generating pressure data 115 d asmaintaining-pressure generating pressure data 116 d. The CPU 113performs the process pertaining to the liquid discharge volume controlusing the pressure sensor-output data 116 a, the calculated-volume-ratiodata 116 b, the injection-pressure generating pressure data 116 c, andthe maintaining-pressure generating pressure data 116 d.

The programs 114 a to 114 g do not need to be stored initially in theROM 114, and for example, the programs can be stored in a portablephysical medium connected to the computer 110, such as a flexible disk(FD), a compact disc-read only memory (CD-ROM), a magneto optical disk(MO), a digital versatile disk (DVD), or an integrated circuit (IC)card, or a fixed physical medium inside or outside the computer 110 suchas HDD. The programs can also be stored in another computer (or aserver) connected to the computer 110 via a public line, the Internet, alocal area network (LAN), or a wide area network (WAN), and downloaded.

As set forth hereinabove, according to an embodiment of the presentinvention, a pressure output to a capillary can be detected withoutinstalling another pressure sensor for detecting the pressure. Thus,facility investment can be reduced can be reduced.

Moreover, volume change in a tube that connects a regulating chamber,valves, a capillary, and the like, can be suppressed. Thus,stabilization of a liquid discharge volume can be achieved.

Although the invention has been described with respect to a specificembodiment for a complete and clear disclosure, the appended claims arenot to be thus limited but are to be construed as embodying allmodifications and alternative constructions that may occur to oneskilled in the art that fairly fall within the basic teaching herein setforth.

1. An injecting apparatus that injects an object into a microbody, theinjecting apparatus comprising: a capillary that injects the object intothe microbody; a regulator that regulates pressure to be applied to theobject; a first valve that is located in a passage between the capillaryand the regulator; a second valve that is located between the firstvalve and the regulator; and a controller that controls the regulator,the first valve, and the second valve, wherein the controller closes thefirst valve in a state where pressure in the capillary is maintained ata first pressure, causes the regulator to generate a second pressure,closes the second valve, opens the first valve to combine the firstpressure with the second pressure to generate a third pressure, appliesthe third pressure to the capillary to cause discharge of the objectfrom the capillary, and causes the regulator to reset the third pressurewhile the object is being discharged from the capillary.
 2. A dischargevolume control device that controls a microinjection apparatus, themicroinjection apparatus including a regulator that regulates andoutputs pressure, a capillary that is filled with an object to bedischarged and connected to the regulator by a tube, a first valve thatis located on regulator side in the tube and a second valve that islocated on capillary side in the tube, and controls volume of the objectdischarged from the capillary, the discharge volume control devicecomprising: an injection pressure regulating unit that opens the secondvalve from a state where the first valve and the second valve are closedto combine an injection-pressure generating pressure with a maintainingpressure to generate an injection pressure, the injection-pressuregenerating pressure being retained between the first valve and thesecond valve to generate the injection pressure that causes the objectto be discharged from the capillary, and the maintaining pressure havingbeen applied to the capillary to prevent backflow of the object into thecapillary, applies the injection pressure to the capillary to causedischarge of the object from the capillary, sets an output pressure ofthe regulator to the injection pressure, and opens the first valve toreapply the injection pressure to the capillary; and a maintainingpressure regulating unit that opens the second valve from a state wherethe first valve and the second valve are closed to combine amaintaining-pressure generating pressure with the injection pressurehaving been applied to the capillary to generate the maintainingpressure, the maintaining-pressure generating pressure being retainedbetween the first valve and the second valve to generate the maintainingpressure, applies the maintaining pressure to the capillary to terminatethe discharge of the object, sets the output pressure of the regulatorto the maintaining pressure, and opens the first valve to reapply themaintaining pressure to the capillary.
 3. A computer-readable recordingmedium that stores therein a computer program for controlling amicroinjection apparatus, the microinjection apparatus including aregulator that regulates and outputs pressure, a capillary that isfilled with an object to be discharged and connected to the regulator bya tube, a first valve that is located on regulator side in the tube anda second valve that is located on capillary side in the tube, andcontrolling volume of the object discharged from the capillary, thecomputer program causing a computer to execute: opening the second valvefrom a state where the first valve and the second valve are closed tocombine an injection-pressure generating pressure with a maintainingpressure to generate an injection pressure, the injection-pressuregenerating pressure being retained between the first valve and thesecond valve to generate the injection pressure that causes the objectto be discharged from the capillary, and the maintaining pressure havingbeen applied to the capillary to prevent backflow of the object into thecapillary; applying the injection pressure to the capillary to causedischarge of the object from the capillary; setting an output pressureof the regulator to the injection pressure; opening the first valve toreapply the injection pressure to the capillary; opening the secondvalve from the state where the first valve and the second valve areclosed to combine a maintaining-pressure generating pressure with theinjection pressure having been applied to the capillary to generate themaintaining pressure, the maintaining-pressure generating pressure beingretained between the first valve and the second valve to generate themaintaining pressure; applying the maintaining pressure to the capillaryto terminate the discharge of the object; setting the output pressure ofthe regulator to the maintaining pressure; and opening the first valveto reapply the maintaining pressure to the capillary.